Structural variability of BM-40/SPARC/osteonectin glycosylation: implications for collagen affinity

Area of Science:

• Biochemistry
• Glycobiology
• Molecular Biology

Background:

• Bone Morphogenetic Protein 40 (BM-40) is a matricellular protein involved in extracellular matrix organization.
• The N-glycosylation of BM-40 can vary depending on its source and cellular environment.
• Understanding these variations is crucial for elucidating BM-40's diverse biological functions.

Purpose of the Study:

• To comprehensively analyze and compare the N-glycan structures of BM-40 isolated from different biological sources.
• To investigate the impact of differential glycosylation on the binding affinity of BM-40 to collagen I.
• To explore the potential functional implications of distinct BM-40 glycoforms in vivo.

Main Methods:

• Purification of BM-40 from human bone, platelets, mouse EHS tumor, and recombinant expression in 293 and osteosarcoma cells.
• Enzymatic digestion (endoglycosidases, exoglycosidases) and High-Performance Liquid Chromatography (HPLC) for N-glycan analysis.
• Surface Plasmon Resonance (SPR) to quantify BM-40 binding to collagen I.

Main Results:

• Bone-derived BM-40 contains high-mannose and biantennary complex N-glycans.
• Platelet and 293 cell-derived BM-40 exclusively show bi- and triantennary complex N-glycans.
• EHS tumor-derived BM-40 exhibits biantennary complex and hybrid structures.
• Recombinant BM-40 from osteosarcoma cells partially recapitulates bone-specific high-mannose glycosylation.
• BM-40 enriched in high-mannose structures demonstrates a higher binding affinity to collagen I.

Conclusions:

• N-glycan structures of BM-40 are source-dependent, with bone BM-40 uniquely possessing high-mannose structures.
• The presence of high-mannose N-glycans on BM-40 significantly enhances its binding affinity to collagen I.
• Differential glycosylation of BM-40 likely contributes to distinct functional roles in various physiological and pathological contexts.